The present invention relates to therapeutic agents for renal diseases and a method for screening and identifying the same. In particular, the present invention relates to a method for screening and identifying such agents, which method was established with close attention directed to the expression of fatty acid-binding protein. The present invention also relates to a cell line established from kidney (proximal renal tubule) cells.
Renal diseases such as nephritis generally presents complex and different pathological aspects, and, when become chronic, may cause serious progression including glomerulosclerosis or interstitial fibrosis, and eventually renal insufficiency. Accordingly, an appropriate treatment in the earlier stage is highly demanded, but there are few therapeutic drugs effective for such purpose. Among the limited number of available drugs, steroids show clear effect but can be accompanied by unacceptably strong adverse side effects. Accordingly, there have been great demands for novel and superior therapeutic agents for renal diseases.
Fatty acid-binding proteins (FABPs) are classified as a group of proteins of molecular weight of around 15 kDa, which present in cytosol and have ability to bind to fatty acids. FABP has been thought to play a role in the regulation of metabolic enzymatic system by transporting and accumulating fatty acids into cells. However, nothing has been known about the correlation between FABP and renal diseases so far.
As for FABP, at least seven molecular species are known, such as liver-type FABP (L-FABP), intestinal-type FABP (I-FABP), heart muscle-type FABP (H-FABP), brain-type FABP (B-FABP), cutaneous/epidermalxe2x80x94type FABP (C-FABP/E-FABP), adipocyte-type FABP (aP2) and peripheral neuron-type FABP (myelin P2). They all have binding activity to fatty acids and considered to be members of a family evolved from a common ancestral gene. Each FABP shows specific histological distribution pattern. The nomenclature of respective FABP means in which organ the FABP was firstly found, but does not necessarily mean that the said FABP exclusively exists in such organ.
In human kidney, at least two FABPs, i.e., liver-type (L-FABP) and heart muscle-type (H-FABP), are expressed and L-FABP is distributed mainly in proximal renal tubule while H-FABP in distal renal tubule (Maatman et al., Biochemical Journal, vol. 288, p. 285-290, 1992; Maatman et al., Biochemical Journal, vol. 273, p. 756-766, 1991).
The expression and distribution of FABP in kidney of rodents is quite different from that of human. In rodents, L-FABP is expressed scarcely in proximal renal tubule and is distributed in distal renal tubule only in a small amount (Maatman et al., Biochemical Journal, vol. 288, p. 285-290, 1992). Predominant FABPs in rodent kidney are H-FABP and kidney-type FABP (K-FABP). H-FABP is mainly distributed in distal renal tubule. K-FABP is thought to be produced as follows. When xcex12U-globulin synthesized in liver is excreted from the circulatory blood to urine in kidney, a portion thereof is reabsorbed by renal tubular cells and converted into K-FABP through the intracellular processing (Kimura et al., FEBS Letters, vol. 246, p. 101-104, 1989).
One of the purposes of the present invention is to provide therapeutic or prophylactic agents for renal disease, which exert effect through a novel mechanism of action, and a method for screening or identifying such drugs. The other purpose of the present invention is to provide a novel cell line useful in the above-mentioned screening or identifying method and the like.
As described above, there had been nothing known about the correlation between FABP and renal diseases before the present inventors found that the decrease in FABP level in urine or renal tissues precedes the infiltration of macrophage or interstitial fibrosis in mouse model of nephritis. The inventors have also found that the L-FABP level in renal tissue is decreased in patients suffering from renal diseases of bad prognosis, and established a method of diagnosing renal diseases based on these findings, which method is focused on FABP (JP-A-11-242026, WO99/27363). The present inventors have continued the research and found that the increase of expression of FABP leads to treatment of renal diseases such as nephritis, in other words, a drug capable of up-regulating the expression of FABP can be a therapeutic agent for renal diseases, and established the present invention.
The present invention provides a method for screening or identifying therapeutic or prophylactic agents for renal diseases, which comprises assaying a test substance for the activity of up-regulating the expression of fatty acid-binding protein (FABP) in animal cells. Further, the present invention provides therapeutic or prophylactic agents selected or identified by such method. Also provided are therapeutic or prophylactic agents for renal diseases comprising, as an active ingredient, an agent having activity of up-regulating the expression of FABP in kidney cells or tissues, for example, a compound having activity of peroxisome proliferator-activated receptor (PPAR) agonist or the like. The present invention also provides a novel mouse proximal renal tubular epithelial cell line useful in the said screening or identifying method.
In patients of renal disease, tissues or cells of kidney are exposed to various kinds of stress such as high proteinuria, ischemia, etc. The therapeutic or prophylactic agents selected or identified by the present method, namely a drug capable of up-regulating the expression of FABP, can protect such cells or tissues. Accordingly, their mechanism of action is based on the protective activity of FABP in kidney tissue or cells (particularly, in renal tubular cells) against stress caused by high proteinuria, ischemia or the like.
Recently, the high proteinuria is regarded as not only a mere indication of renal injury but also one of risky factors per se (Kees-Folts et al., Kidney International, vol. 45, p. 1697-1709, 1994; Eddy et al., Journal of American Society of Nephrology, vol. 5, p. 1273-1287, 1994). It has been known that albumin, a main component of urine, is generally associated with several free fatty acid molecules. Fatty acids binding to urinary proteins are considered to be reabsorbed from brush border membrane of proximal renal tubular epithelial cells, and intracellularly bound to FABP, transported to mitochondria or peroxisome and undergone xcex2-oxidation. In the absence of sufficient amount of FABP, normal xcex2-oxidization of fatty acids can be hindered, which may stimulates the generation of lipid factors (kidney injury factor) that activate macrophages, and lead to the development of interstitial fibrosis via immunological mechanism. In such a case, enhancement of expression of FABP in proximal renal tubular epithelial cells can normalize the fatty acid metabolism and suppress the generation of lipid factor having renal injuring activity and thereby contributing to the improvement of pathology of renal disease.